Environmental tissue injury affects extracellular matrix (ECM) both directly and indirectly: environmental stimuli may directly modify the composition of matrix, e.g. inhaled ozone exposure leads to breakdown of high molecular weight hyaluronan (an abundant ECM component) to low-molecular weight fragments; indirectly, environmental injury induces de-novo production of ECM components or translocation of ECM molecules into the interstitial space, e.g. the serum protein inter-alpha-trypsin inhibitor (IaI) extravasates to the interstitium in fibrotic lung injury. Our research focuses on these two abundant yet understudied molecules, and evaluates how they affect the response to tissue injury. Concretely, our research touches on 3 separate but partially inter-related subjects: 1) To investigate the role of IaI and hyaluronan in airway hyperreactivity after environmental exposures; 2) To investigate the role of IaI and hyaluronan in angiogenesis and tissue healing after injury;and 3) To investigate the role of IaI and hyaluronan in lung transplant rejection. In the first Aim, we were able to show that low-molecular weight hyaluronan is released in the lung airways after ozone exposure in the murine model. Furthermore, we showed that hyaluronan binding through IaI and the cell receptor CD44 is necessary for the mediation of airway hyperreactivity. CD44 is acting in co-receptor fashion with the innate immune receptor TLR4. Finally, hyaluronan binding, or high molecular weight hyaluronan can be used therapeutically to ameliorate airway hyperreactivity in the mouse model. We have identified a number of agents that can effectively inhibit airway hyperresponsiveness in various mouse models of asthma. A patent application is pending and expansion into clinical studies is envisioned. In the second Aim, we investigate the role of IaI and hyaluronan in lung injury. We showed that IaI and hyaluronan are necessary for angiogenesis after lung injury in the mouse model, and that IaI and hyaluronan colocalize in the fibrotic areas of human patients with pulmonary fibrosis, particularly around areas of neovascularization. Furthermore, we were able to show that IaI serum levels in pulmonary fibrosis patients are higher than in control subjects and correlate inversely with gas exchange capacity in these subjects. Interestingly, hypoxia induces these IaI-hyaluronan interactions, a subject that we are investigating currently. Furthermore we identified novel IaI interactions, namely with the ECM molecules complement C3, C4 and vitronectin. These interactions appear to protect against lung inflammation as well as support epithelial wound healing. Other interacting agents have been also identified. IaI therefore emerges as a multipotent "tissue-healing" factor with potential therapeutic applications. Finally, we investigated the effect of a inter-alpha heavy chain, called ITIH4, in inflammation. We have hitherto established that ITIH4 inhibits cell migration, but appears to promote cell activation after endtoxin lung injury. ITIH4 is an acute phase protein which is increased in cancer and COPD patients, so its effects are of particular interest. In the third Aim, we have investigated the pathogenesis of obliterating bronchiolitis, which is the main cause of reejction in lung transplant. Lung transplant rejection is much more common than any of the other solid organs, and we have pursued the hypothesis that this is due to the fact that the lung is in constant contact with the environment, therefore exposed to many immune activating stimuli. We showed that activation of innate immunity through inhaled endotoxin led to alloimmune lung injury in the murine model. Furthermore, we focused on sterile lung injury, which is very common in lung transplant (though pollution, aspiration, ischemia-reperfusion injury, etc.). We were able to show that sterile epithelial injury leads to alloimmune activation and bronchiolitis, specifically through the release of hyaluronan. Furthermore, we showed that endoplsmic reticulum stress is an important mechanism of hyaluronan production in transplant airways injury, and that it appears to promote fibrosis.